A process for preparing a halogen-substituted ribonolactone intermediate that is useful in the synthesis of a 2′-C-methyl-2′-halo nucleoside analogue presents ongoing challenges, particularly where the halogen atom is fluorine.
A key intermediate in the preparation of sugar analogues used in the synthesis of nucleosides and vitamins is 2-C-methyl-D-ribono-lactone. As early as 1880, Scheibler described a process for preparing the lactone (John Sowden, “The Saccharinic Acids” in Adv. Carbohydrate Chem. 12:43-46 (1957), citing C. Scheibler, Berichte 13:2212 (1880)). Unfortunately, product yields were only approximately 10% (Id.). At about the same time, H. Kiliani synthesized 2-methyl-D-ribonolactone by treating D-fructose with calcium hydroxide (H. Kiliani, Berichte, 15:2953 (1882), as cited in F. J. Lopez-Herrera et al., J. Carbohydrate Chemistry, 13(5):767-775 (1994)). However, the process required months to run to completion and product yield was also only approximately 10% (Id. at 768). Kiliani's process, however, enabled him to establish the positions of important functional groups on the compound (John Sowden, “The Saccharinic Acids” in Adv. Carbohydrate Chem. 12:43-46 (1957), citing H. Kiliani, Ann., 213:361 (1883)).
In the early 1960s, Whistler and BeMiller attempted to improve upon Kiliani's synthesis (Roy L. Whistler and J. N. BeMiller, “α-D-Glucosaccharino-1,4-lactone” in Methods in Carbohydrate Chemistry, 2:484-485 (1963)). Whistler and BeMiller added boiling water and calcium hydroxide to D-fructose, flushed the system with nitrogen gas, and repeated the same process. The mixture then was maintained for 6-8 weeks, after which it was treated with CO2 and oxalic acid dihydrate, and filtered under pressure. The residue washed repeatedly to a syrup-like consistency, and filtrates combined; solvent evaporated under reduced pressure and the resultant product allowed to crystallize under refrigeration. The final product yield was still only about 10% (Id. at 485) and the process took two months to complete.
BE 731271 and GB 1189973, assigned to Deutsche Akademie der Wissenchaften, disclosed a process for preparing 3′-fluoronucleosides by reacting a nucleoside with a fluorinating agent such as HF in an organic solvent like THF at temperatures ranging from 130-160° C.
In an attempt to improve product yields, Lopez-Aparicio et al. reported the synthesis of 2-C-methyl-D-ribono-1,4-lactone from 2,3-O-isopropylidene-D-glyceraldehyde as an alternative to the Kiliani synthesis (Lopez-Aparicio et al., Carbohydrate Res., 129:99 (1984), as cited in F. J. Lopez-Herrera et al., J. Carbohydrate Chemistry, 13(5):767-775 (1994) at 768-769). The process of Lopez-Aparicio included condensing 2,3-O-isopropylidene-D-glyceraldehyde with (1-methoxy-carbonyl-ethylidene)triphenylphosphorane to produce methyl E-(S-4,5-dihydroxy-4,5-O-isopropylidene-2-methyl-2-pentenoate; hydrolyzing (in HCl) and photochemically isomerizing the pentenoate; lactonizing the pentenoate product to produce a butenolide; tritylating the butenolide at C-5 by reaction with trityl-chloride and pyridine, followed by cis-hydroxylation with potassium permanganate and methylene chloride in the presence of a crown ether. Final removal of the trityl(triphenylmethyl) group was achieved by reaction with TFA (trifluoroacetic acid) (Id. at 768). Lopez-Aparicio et al. reported product yields of ribonolactone at about 80%, but others were not able to reproduce this figure based on the gram mass amounts of materials provided in the experimental section of their publication. Instead, calculations indicated a percent yield of about 36% ribonolactone. In addition, the process of Lopez-Aparicio et al. was far more complex than the Kiliani synthesis, required the use of toxic reagents such as potassium permanganate and specialized equipment for irradiation to attain photochemical isomerization, and had a minimum of 60 hours reaction time (Id. at 768, 770-772).
None of the foregoing approaches addressed the problem of preparing 2′-C-branched or 2′-disubstituted ribonucleoside analogues.
In 1989, the Asahi Glass Company Ltd. reported the synthesis of fluoronucleosides that had antiviral and antitumor effects (JP 02270864 and JP 01100190). These nucleosides were prepared by treating a 9-(alpha-fluoro-4-beta-hydroxy-1-beta-cyclopentyl)pyrimidine derivative with trifluoromethanesulphonyl chloride, p-toluenesulphonyl chloride, methanesulphonyl chloride or imidazolylsulphonyl chloride in the presence of a base, followed by reduction (JP 02270864). In a second synthetic method, 2′,3′-deoxy-2′,3′-didehydro-2′-fluoronucleosides were obtained by the dehydrogenation of a 2′-deoxy-2′-fluororibofuranosyl derivative, or by dehydrogenation of a 2′,3′-dideoxy-2′-fluoro-3′-halo-ribonucleoside derivative (JP 01100190).
In 1990, Bobek et al. disclosed the synthesis of antiviral, antitumor, and antimicrobial arabinopyranosyl nucleoside derivatives that had a fluorine atom at the 2′-position of the pyranose ring (U.S. Pat. No. 4,918,056). These compounds were prepared by the condensation of a pyrimidine, purine, or 1,3-oxazine nucleobase with an hydroxyl group-blocked, acylated 2-deoxy-2,2-difluoro-D-arabinopyranoside and/or an acylated 2-deoxy-2-bromo-2-fluoro-D-arabinopyranoside (Id.).
In 1997 Harry-O'Kuru et al. described a synthetic route for preparing 2′-C-branched ribonucleosides (Harry-O'Kuru et al., J. Org. Chem., 62:1754-9 (1997)). Commercially available 1,3,5-tri-O-benzoyl-α-D-ribofuranose was used as the starting material, which was prepared from D-ribose or D-arabinose (D-arabinopyranose). The 1,3,5-tri-O-benzoyl-α-D-ribofuranose was oxidized at the free 2-OH with Dess-Martin periodinane reagent, and produced 1,3,5-tri-O-benzoyl-2-keto-ribofuranose as well as its corresponding hydrate. The desired product and hydrate were stirred with excess MgSO4 and permitted to stand overnight. The mixture was then filtered and concentrated in order to produce a substantially pure ketone product. The resultant 2-ketosugar was treated with MeMgBr/TiCl4 (or alternatively with MeTiCl3, CH2═CHMgBr/CeCl3, or TMSC≡CLi/CeCl3), which produced an anomeric mixture of the desired 1,3,5-tri-O-benzoyl-2-substituted alkyl-, alkenyl- or alkynyl-ribofuranoside and its transesterified isomers, α- and β-2,3,5-tri-O-benzoyl-2-substituted alkyl, alkenyl or alkynyl ribofuranoside in a nearly 5:3 ratio of desired product to isomeric forms (Id. at 1755). The 2-alkylated ribofuranosides then were converted to a single, desired product, 1,2,3,5-tetrabenzoyl-2-alkylribofuranoside, by treatment with benzoyl chloride, DMAP and triethylamine in a reported approximately 70% yield with a β/α ratio of 4:1 (Id.).
In 1998, Chambers et al. reported the synthesis of 2′,3′-dideoxy-3′-fluorouridine compounds by reaction of a corresponding anhydronucleoside with hydrogen fluoride in the presence of an organo-iron compound and in an organic solvent (U.S. Pat. No. 5,717,086).
Recent reports of syntheses of 2′ and/or 3′ halonucleosides have been disclosed by Pharmasset, Inc., The University of Georgia Research Foundation, Inc., and Emory University.
WO 05/003147 (also US 2005/0009737) to Pharmasset, Inc., described the synthesis of 2′-C-methyl-2′-fluoro nucleoside analogues by one of two general synthetic routes: alkylating an appropriately modified carbohydrate compound, fluorinating it, and then coupling it to a desired nucleobase, or glycosylating a desired nucleobase to form a nucleoside, then alkylating the nucleoside, and finally fluorinating the preformed nucleoside. Pharmasset's first approach utilized a modified carbohydrate that was an hydroxyl group-protected lactone, which was alkylated with a reagent such as methyl lithium in an anhydrous solvent like THF, and then was reacted with a commercially available fluorinating agent like DAST or Deoxofluor, followed by a deprotection step. The reaction proceeded with inversion at the 2′-position such that the fluorine atom was in the “down” or ribo configuration. Pharmasset's second synthetic route comprised the modification of a commercially available nucleoside whose hydroxyl groups were protected by protective groups known in the art. The nucleoside was oxidized at the 2′-position to provide a 2′-ketone, and the 2′-ketone was reacted with an alkylating agent such as methyl lithium in THF at about 0° C. to afford a 2′(S) or 2′-methyl “down”, 2′-hydroxyl “up” configured nucleoside tertiary alcohol. A fluorine atom then was introduced by reacting the nucleoside with a commercially available fluorinating reagent such as DAST in an anhydrous, aprotic solvent like toluene with inversion at the 2′-position to afford a 2′-C-methyl “up”, 2′-fluoro “down” configuration of the nucleoside. However, by either synthetic route, Pharmasset's isolation and purification methods were impractical/inefficient and product yield was very low in all examples provided.
PCT Publication No. WO 2006/031725 to Pharmasset, Inc. describes the synthesis of 2′-C-substituted-2′-deoxy-2′-halo nucleosides via the nucleophilic ring opening of a 5-membered ring cyclic sulfate intermediate derived from 4,5-di-O-protected-2-methyl-2,3-dihydroxy-pentanoic acid with fluoride to produce a 2-methyl-2-fluoro 4,5-di-O-protected fluorinated acyclic sulfate ester compound. The fluorinated sulfate ester is treated with acid to deprotect the 4,5-hydroxyl groups and cyclized to 2′-fluoro-2′-C-methyl-γ-ribonolactone. The ribonolactone is then converted to the 2′-C-methyl-2′-deoxy-2′-halo nucleosides by reduction of the lactone and coupling with an appropriate base.
WO 2006/012440 to Pharmasset, Inc. describes the synthesis of 2′-C-substituted-2′-deoxy-2′-halo nucleosides via a 2′-fluoro-2′-C-substituted-γ-ribonolactone intermediate. The 2′-fluoro-2′-C-substituted-γ-ribonolactone is formed by cyclization of a 2-fluoro-4,5-di-O-protected-2,3-dihydroxy-pentanoic acid ethyl ester intermediate upon treatment with acid. The fluorination reaction is achieved by treating 4,5-di-O-protected-2-hydroxy-3-O-protected-pentanoic acid ethyl ester with DAST.
Otto et al. reported the synthesis and use of beta-2′ or beta-3′-halonucleosides for the treatment of HIV, hepatitis B, and undesired cellular proliferation (U.S. Pat. No. 6,949,522). The syntheses disclosed produced 2′,3′-dideoxy, 3′,3′-dihalo nucleosides from glyceraldehyde or a sugar ring starting material; 2′,3′-dideoxy, 3′-halo nucleosides from a lactol starting material; and 2′,3′-dideoxy-2′-halo nucleosides from glyceraldehyde as a starting material that proceeds via a lactone intermediate that is selectively reduced to afford a 2′,3′-hydro product.
Clark et al. disclosed the synthesis and antiviral activity of 2′-deoxy-2′-fluoro-2′-C-methylcytidine as an inhibitor of hepatitis C virus (Clark et al., J. Med. Chem. 2005, 48:5504-5508). Synthesis of the product compound proceeded through N4-benzoyl-1-(2-methyl-3,5-di-O-benzoyl-β-D-arabinofuranosyl)cytosine as a key intermediate, which was oxidized to the corresponding 2′-ketone derivative by reaction with trifluoroacetic anhydride in DMSO under Swern oxidation conditions. The 2′-ketone derivative was reacted with methyllithium at −78° C. in diethyl ether to afford protected 1-[2-C-methyl-3,5-O-(tetraisopropyldisiloxane-1,3-diyl)1-β-D-arabinofuranosyl]cytosine, and the 3′,5′-silyl protecting group was removed by reaction in TBAF/acetic acid. Clark et al. warn against the use of DAST for fluorination of tertiary alcohol groups because the reaction is substrate specific and stereochemically unpredictable.
Shi et al. reported the syntheses and antiviral activities of a series of D- and L-2′-deoxy-2′-fluororibonucleosides in a hepatitis C replicon system (Shi et al., Bioorganic & Medicinal Chemistry (2005), 13:1641-1652). The halo-substituted nucleosides tested had a single halo substituent at the 2′-position on the nucleoside sugar, and were prepared by direct conversion of D-2,2′-anhydrocytidine to (2′R)-D-2′-deoxy-2′-fluorocytidine by reaction with potassium fluoride and crown ether according to the method of Mengel and Guschlbauer (Angew. Chem. Int. Ed. Engl. 1978, 17:525).
There remains a need for discovering improved synthetic routes and new synthetic intermediates in the preparation of 2′-C-methyl-2′-halo-nucleoside analogue derivatives.
It is an object of the present invention to provide a stereochemically predictable and reliable process for the selective addition of alkyl and halo substituents at the 2′-C-position of a nucleoside analogue.
It is another object of the present invention to provide an efficient process that utilizes a minimum number of steps and a readily available, inexpensive starting material for preparing a key intermediate in the synthesis of a 2′-C-methyl-2′-halo-nucleoside analogue.
It is still another object of the present invention to provide a process that employs non-toxic reagents and provides the key intermediate in good percent product yield.